A new Drosophila melanogaster research resource: CRISPR-induced mutations for clonal analysis of fourth chromosome genes.

As part of an ongoing effort to generate comprehensive resources for the experimental analysis of fourth chromosome genes in Drosophila melanogaster, the Fourth Chromosome Resource Project has used CRISPR mutagenesis with single guide RNAs to isolate mutations in 62 of the 80 fourth chromosome, protein-coding genes. These mutations were induced on a fourth chromosome bearing a basal FRT insertion to facilitate experimental approaches involving FLP recombinase-induced mitotic recombination. To permit straightforward comparisons among mutant stocks, most of the mutations were generated on isogenic fourth chromosomes, which were then crossed into a common genetic background.

Control of fate specification within the dorsal head of Drosophila melanogaster.

During development, unique combinations of transcription factors and signaling pathways carve the nascent eye-antennal disc of the fruit fly Drosophila melanogaster into several territories that will eventually develop into the compound eye, ocelli, head epidermis, bristles, antenna and maxillary palpus of the adult head. Juxtaposed patterns of Hedgehog (Hh) and Decapentaplegic (Dpp) initiate compound eye development, while reciprocal domains of Dpp and Wingless (Wg) induce formation of the antennal and maxillary palp fields. Hh and Wg signaling, but not Dpp, contribute to the patterning of the dorsal head vertex.

Fourth Chromosome Resource Project: a comprehensive resource for genetic analysis in Drosophila that includes humanized stocks.

The fourth chromosome is the final frontier for genetic analysis in Drosophila. Small, heterochromatic, and devoid of recombination the fourth has long been ignored. Nevertheless, its long arm contains 79 protein-coding genes.

Polycomb safeguards imaginal disc specification through control of the Vestigial-Scalloped complex.

A fundamental goal of developmental biology is to understand how cell and tissue fates are specified. The imaginal discs of Drosophila are excellent model systems for addressing this paradigm as their fate can be redirected when discs regenerate after injury or when key selector genes are misregulated. Here, we show that when Polycomb expression is reduced, the wing selector gene vestigial is ectopically activated.

Polycomb safeguards imaginal disc specification through control of the Vestigial-Scalloped complex.

Unlabelled: A fundamental goal of developmental biology is to understand how cell and tissue fates are specified. The imaginal discs of are excellent model systems for addressing this paradigm as their fate can be redirected when discs regenerate after injury or when key selector genes are mis-regulated. Here, we show that when expression is reduced, the wing selector gene is ectopically activated.

A CUT&RUN protocol to determine patterns of epigenetic marks in imaginal discs of Drosophila.

Cleavage Under Targets & Release Using Nucleases (CUT&RUN) sequencing is a technique used to study gene regulation. The protocol presented here has been used successfully to identify the pattern of histone modifications within the genome of the eye-antennal disc of the fruit fly, Drosophila melanogaster. In its present form, it can be used to analyze genomic features of other imaginal discs.

The early history of the eye-antennal disc of Drosophila melanogaster.

A pair of eye-antennal imaginal discs give rise to nearly all external structures of the adult Drosophila head including the compound eyes, ocelli, antennae, maxillary palps, head epidermis, and bristles. In the earliest days of Drosophila research, investigators would examine thousands of adult flies in search of viable mutants whose appearance deviated from the norm. The compound eyes are dispensable for viability and perturbations to their structure are easy to detect.

A multi-gene knockdown approach reveals a new role for Pax6 in controlling organ number in Drosophila.

Genetic screens are designed to target individual genes for the practical reason of establishing a clear association between a mutant phenotype and a single genetic locus. This allows for a developmental or physiological role to be assigned to the wild-type gene. We previously observed that the concurrent loss of Pax6 and Polycomb epigenetic repressors in Drosophila leads the eye to transform into a wing.

Retinal Expression of the Drosophila eyes absent Gene Is Controlled by Several Cooperatively Acting Cis-regulatory Elements.

The eyes absent (eya) gene of the fruit fly, Drosophila melanogaster, is a member of an evolutionarily conserved gene regulatory network that controls eye formation in all seeing animals. The loss of eya leads to the complete elimination of the compound eye while forced expression of eya in non-retinal tissues is sufficient to induce ectopic eye formation. Within the developing retina eya is expressed in a dynamic pattern and is involved in tissue specification/determination, cell proliferation, apoptosis, and cell fate choice.

The Drosophila Wilms׳ Tumor 1-Associating Protein (WTAP) homolog is required for eye development.

Sine Oculis (So), the founding member of the SIX family of homeobox transcription factors, binds to sequence specific DNA elements and regulates transcription of downstream target genes. It does so, in part, through the formation of distinct biochemical complexes with Eyes Absent (Eya) and Groucho (Gro). While these complexes play significant roles during development, they do not account for all So-dependent activities in Drosophila.

Dual transcriptional activities of SIX proteins define their roles in normal and ectopic eye development.

The SIX family of homeodomain-containing DNA-binding proteins play crucial roles in both Drosophila and vertebrate retinal specification. In flies, three such family members exist, but only two, Sine oculis (So) and Optix, are expressed and function within the eye. In vertebrates, the homologs of Optix (Six3 and Six6) and probably So (Six1 and Six2) are also required for proper eye formation.

A dissection of the teashirt and tiptop genes reveals a novel mechanism for regulating transcription factor activity.

In the Drosophila eye the retinal determination (RD) network controls both tissue specification and cell proliferation. Mutations in network members result in severe reductions in the size of the eye primordium and the transformation of the eye field into head cuticle. The zinc-finger transcription factor Teashirt (Tsh) plays a role in promoting cell proliferation in the anterior most portions of the eye field as well as in inducing ectopic eye formation in forced expression assays.

A novel system for the launch of alphavirus RNA synthesis reveals a role for the Imd pathway in arthropod antiviral response.

Alphaviruses are RNA viruses transmitted between vertebrate hosts by arthropod vectors, primarily mosquitoes. How arthropods counteract alphaviruses or viruses per se is not very well understood. Drosophila melanogaster is a powerful model system for studying innate immunity against bacterial and fungal infections.

The non-conserved C-terminal segments of Sine Oculis Homeobox (SIX) proteins confer functional specificity.

The Sine Oculis Homeobox (SIX) proteins play critical roles in organogenesis and are defined by the presence of two evolutionarily conserved functional motifs: a homeobox DNA binding domain and the SIX protein-protein interaction domain. Members of this transcription factor family can be divided into three subgroups: Six1/2, Six4/5, and Six3/6. This partitioning is based mainly on protein sequence similarity and genomic architecture, and not on specificities of DNA binding or binding partners.

Transcriptional activities of the Pax6 gene eyeless regulate tissue specificity of ectopic eye formation in Drosophila.

Pax genes encode DNA binding proteins that play pivotal roles in the determination of complex tissues. Members of one subclass, Pax6, function as selector genes and play key roles in the retinal development of all seeing animals. Mutations within the Pax6 homologs including fly eyeless, mouse Small eye and human Pax6 lead to severe retinal defects in their respective systems.

Differential requirements for the Pax6(5a) genes eyegone and twin of eyegone during eye development in Drosophila.

In eye development the tasks of tissue specification and cell proliferation are regulated, in part, by the Pax6 and Pax6(5a) proteins respectively. In vertebrates, Pax6(5a) is generated as an alternately spliced isoform of Pax6. This stands in contrast to the fruit fly, Drosophila melanogaster, which has two Pax6(5a) homologs that are encoded by the eyegone and twin of eyegone genes.

Sine oculis, a member of the SIX family of transcription factors, directs eye formation.

The initiation of eye formation in all seeing animals is controlled by a group of selector genes that together forms the retinal determination cascade. In Drosophila, mice and humans, loss-of-function mutations lead to defects in eye and/or head development. While ectopic expression of these genes is sufficient to direct non-retinal tissues towards an eye fate, the ability of each gene to initiate eye formation is neither unlimited nor equal.

Conservation of an inhibitor of the epidermal growth factor receptor, Kekkon1, in dipterans.

Regulation of epidermal growth factor receptor (EGFR) signaling requires the concerted action of both positive and negative factors. While the existence of numerous molecules that stimulate EGFR activity has been well documented, direct biological inhibitors appear to be more limited in number and phylogenetic distribution. Kekkon1 (Kek1) represents one such inhibitor.

The Eye Specification Network in Drosophila.

One of the most exciting revelations in retinal biology is the realization that the molecules and mechanisms that regulate eye development have been conserved in all seeing animals including such diverse organisms as the fruit fly, mouse and man. The emerging commonality among mechanisms used in eye development allows for the use of model systems such as the fruit fly, Drosophila melanogaster, to provide key insights into the development and diseases of the mammalian eye. Eye specification in Drosophila is controlled, in part, by the concerted activities of eight nuclear proteins and several signal transduction cascades that together form a tightly woven regulatory network.